There are two distinct TNF-specific plasma membrane-localized receptors, type I 55 kDa TNFR (TNFR1) and type II 75 kDa TNFR (TNFR2). TNFR1 is expressed ubiquitously, whereas TNFR2 is tightly regulated and found predominantly on endothelial cells (EC). TNF via TNFR1 and associated adaptor molecules elicits a broad spectrum of biological effects including proliferation, differentiation and apoptosis. Little is known regarding the proteins recruited to TNFR2 and the function of TNFR2 in EC. Anti-TNF therapy targeting both TNFR1 and TNFR2 has been successful in treating rheumatoid arthritis but failed in the treatment of cardiovascular diseases. We hypothesize that TNFR2 signaling plays beneficial roles in the cardiovascular system such as ischemia-initiated arteriogenesis/angiogenesis and remodeling. We show that genetic loss of TNFR2, but not of TNFR1, impairs ischemia initiated blood flow recovery resulting in critical limb ischemia. This may occur via impaired arteriogenesis, angiogenesis or mobilization of endothelial progenitor cells. We have established various assays in order to dissect these defects in TNFR2-KO mice. Meanwhile, we have made efforts to define the critical downstream effectors in TNFR2 angiogenic signaling. We show that TRAF2, the only previously known TNFR2-interacting protein, regulates the anti-apoptotic function of TNFR2;we have identified the first TNFR2-specific kinase, Etk, critical for TNF-induced EC migration and tube formation. Thus TNF activation of TNFR2 leads to distinct but cooperative downstream signaling of EC survival, migration and tube formation during arteriogenesis/angiogenesis. The successful breeding of genetically-deficient mice for TNFR1, TNFR2, TRAF2 and Etk, isolation of mouse muscle microvessel EC from these mice and establishment of in vivo angiogenesis/arteriogenesis assays will allow us to dissect the signaling by TNFR2. We propose the following specific aims: 1) Define the mechanism by which TNFR2 mediates angiogenesis/arteriogenesis using TNFR2-KO mice. 2) Dissect TNFR2 signaling in EC. 3) Define the role of EC-expressed TNFR2 in inflammatory angiogenesis using transgenic mouse models. This proposal should provide novel information on the role of TNFR2 and its downstream effectors TRAF2 and Etk in inflammatory angiogenesis/arteriogenesis, and facilitate development of new therapeutic approaches to control angiogenesis/arteriogenesis-dependent cardiovascular diseases. Growth and maturation of blood vessels are critical for the repair of ischemic heart, brain and extremity. We will study the role of a membrane receptor TNFR2 in the process. Our work may lead to better treatments for ischemic diseases.